The recent call for elimination of malaria caused by Plasmodium falciparum (Pf) and eventual eradication of all malaria has focused attention on this disease, which is responsible for hundreds of millions of cases and a million deaths annually. An ideal tool for eliminating Pf, the causative agent of 99% of all malaria deaths, would be a highly effective vaccine that prevents blood stage infection and thereby prevents all disease and transmission. When attenuated Pf sporozoites (PfSPZ) are administered by the bite of infected mosquitoes, >90% of human volunteers are protected against experimental Pf challenge and protection lasts at least 10 months. Sanaria's goal is to develop and commercialize an attenuated PfSPZ vaccine that prevents Pf blood stage infection in >90% of recipients;a vaccine that could be used to eliminate Pf from the world. This vaccine has the potential for >$1 billion annual revenues in markets in the developed and developing world. Sanaria has succeeded in establishing robust, reproducible, and consistent manufacture and release of clinical lots of its PfSPZ Vaccine, received FDA approval to proceed with clinical trials (IND approval), and in the 2nd quarter of 2009 will be initiating a Phase 1 clinical trial to assess safety, immunogenicity, and protective efficacy of the PfSPZ Vaccine. After demonstrating safety in that trial, the goal is to move as swiftly as possible to safety and proof of concept efficacy studies in African adults, young children, and infants. These will be followed by additional Phase 2 studies, including dose optimization studies, and then pivotal Phase 3 studies to support licensure. There is a clearcut path to taking such a mosquito-produced vaccine forward to registration and commercialization. However, it would be ideal if the sporozoites could be produced in culture instead of in mosquitoes. After a mosquito ingests malaria parasites called gametocytes when feeding on humans, the parasite develops in the mosquito from the gametoctyes into gametes, zygotes, ookinetes, and finally oocysts containing sporozoites. Pf oocysts containing sporozoites were produced in culture in 1993, but the finding was never replicated, primarily because it was not possible to reproducibly produce Pf ookinetes in culture. Infective P. berghei sporozoites have been produced in culture. It is the goal of this project to establish a method of producing in culture Pf sporozoites that can be used in an attenuated sporozoite vaccine. This method of production would eliminate the need for producing sporozoites in mosquitoes. The major impediment to utilizing the system established in 2002 for producing P. berghei sporozoites in vitro to produce Pf sporozoites in vitro has been the capacity to produce large quantities of viable Pf ookinetes. This limitation has now been overcome. In the 1st 6 months of this project we will optimize ookinete production and purification. Subsequently we will use Pf ookinetes to optimize production of Pf sporozoites in vitro, and demonstrate infectivity of these sporozoites to human hepatocytes in vitro. The long term goal is to deliver a method to produce in vitro Pf sporozoites for use in an attenuated Pf sporozoite vaccine. PUBLIC HEALTH RELEVANCE: A highly effective malaria vaccine would have an enormous public health benefit. Sanaria's attenuated malaria sporozoite vaccine (PfSPZ Vaccine) is expected to be highly effective and will be studied in clinical trials in 2009. The PfSPZ Vaccine is manufactured in mosquitoes. This research aims to eliminate the need for mosquitoes in manufacturing the PfSPZ Vaccine.